WO1998037027A1 - Method for acquiring grain-shaped growth of a microorganism in a reactor - Google Patents

Method for acquiring grain-shaped growth of a microorganism in a reactor Download PDF

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Publication number
WO1998037027A1
WO1998037027A1 PCT/NL1998/000100 NL9800100W WO9837027A1 WO 1998037027 A1 WO1998037027 A1 WO 1998037027A1 NL 9800100 W NL9800100 W NL 9800100W WO 9837027 A1 WO9837027 A1 WO 9837027A1
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WO
WIPO (PCT)
Prior art keywords
reactor
microorganism
liquid
phase
during
Prior art date
Application number
PCT/NL1998/000100
Other languages
French (fr)
Inventor
Joseph Johannes Heijnen
Marinus Cornelis Maria Van Loosdrecht
Original Assignee
Technische Universiteit Delft
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Filing date
Publication date
Application filed by Technische Universiteit Delft filed Critical Technische Universiteit Delft
Priority to EP98905876A priority Critical patent/EP0964831B1/en
Priority to US09/367,985 priority patent/US6566119B1/en
Priority to DE1998607659 priority patent/DE69807659T2/en
Priority to AT98905876T priority patent/ATE223356T1/en
Priority to DK98905876T priority patent/DK0964831T3/en
Publication of WO1998037027A1 publication Critical patent/WO1998037027A1/en

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    • C—CHEMISTRY; METALLURGY
    • C02—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
    • C02F—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
    • C02F3/00—Biological treatment of water, waste water, or sewage
    • C02F3/02—Aerobic processes
    • C02F3/12—Activated sludge processes
    • C02F3/1236—Particular type of activated sludge installations
    • C02F3/1263—Sequencing batch reactors [SBR]
    • C—CHEMISTRY; METALLURGY
    • C02—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
    • C02F—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
    • C02F3/00—Biological treatment of water, waste water, or sewage
    • C02F3/02—Aerobic processes
    • C02F3/12—Activated sludge processes
    • C02F3/20—Activated sludge processes using diffusers
    • C—CHEMISTRY; METALLURGY
    • C02—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
    • C02F—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
    • C02F3/00—Biological treatment of water, waste water, or sewage
    • C02F3/30—Aerobic and anaerobic processes
    • C02F3/302—Nitrification and denitrification treatment
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
    • C12N1/02—Separating microorganisms from their culture media
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
    • C12N1/20—Bacteria; Culture media therefor
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
    • C12N1/36—Adaptation or attenuation of cells
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
    • Y02W—CLIMATE CHANGE MITIGATION TECHNOLOGIES RELATED TO WASTEWATER TREATMENT OR WASTE MANAGEMENT
    • Y02W10/00—Technologies for wastewater treatment
    • Y02W10/10—Biological treatment of water, waste water, or sewage
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
    • Y10S—TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10S435/00—Chemistry: molecular biology and microbiology
    • Y10S435/8215—Microorganisms
    • Y10S435/822—Microorganisms using bacteria or actinomycetales
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
    • Y10S—TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10S435/00—Chemistry: molecular biology and microbiology
    • Y10S435/8215—Microorganisms
    • Y10S435/911—Microorganisms using fungi

Definitions

  • the present invention relates to a method of acquiring granular growth of a microorganism in a reactor containing a liquid phase which comprises a substrate, wherein in a first step said substrate is converted by the microorganism resulting in the formation of and growth on a phase comprising the organism while the liquid phase is being mixed, in a second step mixing in the reactor is stopped to allow part of the solid phase to settle, and in a third step the reactor is partly emptied by discharging the top part of the reactor contents, which reactor is subsequently replenished with substrate-comprising liquid, to repeat steps 1 to 3.
  • the method according to the invention is therefore characterized in that the microorganism is an aerobic microorganism, in that at least during the first step a third phase is present, which third phase comprises oxygen-containing gas being fed to the reactor during the first step while the contents of the reactor are kept in turbulence, and in that settling occurs in the second step and takes less time than the height of liquid in the reactor at the end of the first step divided by a settling velocity of at least 5 metres per hour.
  • the compound is fed to the reactor in pulses.
  • This causes the organisms in a granule to be flooded with compound.
  • the organisms at the outside of the granule are unable to process such a supply of compound, the compound gets the opportunity to diffuse into the interior of the granule. This is especially important if the compound to be converted is a nutrient .
  • An example of this is carbohydrate fermentation for the preparation of lactic acid.
  • the contents of the reactor are substantially continuously mixed during the first step.
  • Sung et al. prefer periodical mixing during which only slight shearing forces occur.
  • continuous turbulent mixing subjects floes to mixing forces, allowing them to be discharged more easily in the third step.
  • the organisms in the reactor form into granules more quickly.
  • Turbulent mixing is conveniently carried out by feeding oxygen-containing gas into, for instance, an airlift-reactor or bubbling-bed reactor.
  • An interesting application of the method according to the invention is characterized in that the conversion is a nitrification-denitrification-conversion in which the oxygen-containing gas is only supplied during the first part of the first step for the completion of the nitrification, and that the reactor operates during the remainder of the first step under substantially anaerobic conditions for the completion of the denitrification. If desired, the gas may during this first step be recirculated over the reactor. Due to recirculation all oxygen is used up and turbulence is maintained.
  • the organism-comprising granules must be present, or at least conditions promoting the formation of granules must be provided before starting up the reactor. It is useful, for instance, to feed the reactor with carrier particles to which organisms adhere, or are able to adhere. It has been shown that a mycelium-forming fungus can also be used as carrier.
  • settling occurs in the second step, taking less time than the height of liquid in the reactor at the end of the first step divided by a settling velocity of at least 10 metres per hour, preferably at least 15 metres per hour.
  • a clarification step taking characteristically 10 to 30 minutes.
  • the settling velocity is then only 1 metre per hour, and applied to aerobic organisms such a method will not result in granulation.
  • the clarification step merely serves to separate organisms and treated water, is also the clarification step in the method according to the invention of essential importance for the induction of granulation.
  • Fig.- 1 is a graphical representation of the carbon dioxide percentage in gas discharged from a bubbling-bed reactor during a cycle of the method according to the invention.
  • Fig. 2 is an illustration of a granule composed of aerobic organisms.
  • Example l is an illustration of a granule composed of aerobic organisms.
  • a bubbling-bed reactor (2.5 litres; height/diameter 20) was fed with 1.25 litres of a solution serving as model for waste water, comprising 8.7 mM ethanol, 5 mM ammonium chloride, 4.7 mM potassium phosphate, 2.4 mM magnesium sulphate, 0.48 calcium chloride and per litre solution 1.5 ml of a standard solution of trace elements.
  • the solution was inoculated with aerobically active sludge from a water treatment plant .
  • the model waste water in the bubbling bed reactor was subjected to a cyclic treatment at pH 6-8 and a temperature of 20°C. The treatment consisted of i) aerating for 4 hours at a flow rate of 1.5 litres air per minute (Fig.
  • l shows the carbon dioxide percentage in the gas discharged from a bubbling-bed reac- tor during this phase. This percentage is a measure of the conversion of the ethanol) , ii) the one-minute stoppage of aeration, and iii) draining model waste water from the bubbling-bed reactor at the half-way point of the column of liquid. Any biomass present during draining in the top half of the solution, was discharged together with the effluent. Finally, iv) the bubbling-bed reactor was replenished with a volume of model waste water equal to that of the discharged effluent . The cycle was then resumed with four hours aeration of the solution.
  • Fig. 2 shows the granules comprised of aerobic microorganisms, obtained by the method according to the invention. The average size is 3 mm.

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  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Organic Chemistry (AREA)
  • Health & Medical Sciences (AREA)
  • Genetics & Genomics (AREA)
  • Biotechnology (AREA)
  • Zoology (AREA)
  • Wood Science & Technology (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Microbiology (AREA)
  • Tropical Medicine & Parasitology (AREA)
  • Hydrology & Water Resources (AREA)
  • Virology (AREA)
  • Medicinal Chemistry (AREA)
  • Biochemistry (AREA)
  • General Engineering & Computer Science (AREA)
  • General Health & Medical Sciences (AREA)
  • Water Supply & Treatment (AREA)
  • Biodiversity & Conservation Biology (AREA)
  • Biomedical Technology (AREA)
  • Environmental & Geological Engineering (AREA)
  • Cell Biology (AREA)
  • Purification Treatments By Anaerobic Or Anaerobic And Aerobic Bacteria Or Animals (AREA)
  • Micro-Organisms Or Cultivation Processes Thereof (AREA)
  • Cereal-Derived Products (AREA)
  • Apparatus Associated With Microorganisms And Enzymes (AREA)
  • Biological Treatment Of Waste Water (AREA)

Abstract

The invention relates to a method of acquiring granular growth of a microorganism in a reactor containing a liquid medium. Surprisingly, according to the invention, aerobic microorganisms also can be induced to granular growth by maintaining specific culture conditions. During a first step an oxygen-containing gas is supplied and the reactor contents are kept in turbulence. In a second step, after a short settling period, the top part of the reactor medium is discharged.

Description

Method for acquiring grain-shaped growth of a microorganism in a reactor
The present invention relates to a method of acquiring granular growth of a microorganism in a reactor containing a liquid phase which comprises a substrate, wherein in a first step said substrate is converted by the microorganism resulting in the formation of and growth on a phase comprising the organism while the liquid phase is being mixed, in a second step mixing in the reactor is stopped to allow part of the solid phase to settle, and in a third step the reactor is partly emptied by discharging the top part of the reactor contents, which reactor is subsequently replenished with substrate-comprising liquid, to repeat steps 1 to 3.
Such a method has been described by Sung S . et al . (Laboratory studies of the anaerobic sequencing batch reactor, in Water Environment Research, 61. (3) , p. 294, 1995) . In this method an anaerobic conversion is carried out, the contents of the reactor are then clarified in characteristically 10-30 minutes after which the top part of the reactor contents is discharged. According to this publication work with anaerobically activated sludge has long been known, although it was not recognized in the beginning (1966) that what was occurring was "granulation" of the biomass. Granulation under methanogenic conditions (that is to say in the absence of oxygen) is often explained by the specific need of the respective organisms to exchange substrates the so-called "interspecies hydrogen transfer", or to reduce the toxicity of oxygen.
Surprisingly, applicant has found that granulation can also occur under aerobic and turbulent conditions. The method according to the invention is therefore characterized in that the microorganism is an aerobic microorganism, in that at least during the first step a third phase is present, which third phase comprises oxygen-containing gas being fed to the reactor during the first step while the contents of the reactor are kept in turbulence, and in that settling occurs in the second step and takes less time than the height of liquid in the reactor at the end of the first step divided by a settling velocity of at least 5 metres per hour. • The formation of granules under turbulent aerobic conditions is unexpected, because organisms in a granule are under very great stress as due to the size of the granule, nutrients have to diffuse over large distances before reaching the interior of a granule . The fact that in addition aerobic organisms require oxygen, increases the stress still more so that one would expect such aerobic organisms only to flocculate. In addition, the expert would expect the granules to disintegrate due to the great shearing forces caused by the turbulence . In the present application an aerobic microorganism is understood to be both an obligate and a facultative aerobic microorganism.
Preferably the compound is fed to the reactor in pulses. This causes the organisms in a granule to be flooded with compound. As the organisms at the outside of the granule are unable to process such a supply of compound, the compound gets the opportunity to diffuse into the interior of the granule. This is especially important if the compound to be converted is a nutrient . An example of this is carbohydrate fermentation for the preparation of lactic acid.
In accordance with a favourable embodiment the contents of the reactor are substantially continuously mixed during the first step. To promote the formation of aggregates, Sung et al. prefer periodical mixing during which only slight shearing forces occur. According to the invention, however, continuous turbulent mixing subjects floes to mixing forces, allowing them to be discharged more easily in the third step. In addition, applicant has established by experimentation that if microorganisms are subjected to great shearing forces, the result is a more robust granule. Thus according to the invention the organisms in the reactor form into granules more quickly. Turbulent mixing is conveniently carried out by feeding oxygen-containing gas into, for instance, an airlift-reactor or bubbling-bed reactor.
An interesting application of the method according to the invention is characterized in that the conversion is a nitrification-denitrification-conversion in which the oxygen-containing gas is only supplied during the first part of the first step for the completion of the nitrification, and that the reactor operates during the remainder of the first step under substantially anaerobic conditions for the completion of the denitrification. If desired, the gas may during this first step be recirculated over the reactor. Due to recirculation all oxygen is used up and turbulence is maintained. When applying the method according to the invention, the organism-comprising granules must be present, or at least conditions promoting the formation of granules must be provided before starting up the reactor. It is useful, for instance, to feed the reactor with carrier particles to which organisms adhere, or are able to adhere. It has been shown that a mycelium-forming fungus can also be used as carrier.
According to a very favourable embodiment settling occurs in the second step, taking less time than the height of liquid in the reactor at the end of the first step divided by a settling velocity of at least 10 metres per hour, preferably at least 15 metres per hour. Thus the presence of granules in the reactor is strongly favoured in comparison with the presence of floes. As mentioned above, Sung et al . describe a clarification step taking characteristically 10 to 30 minutes. The settling velocity is then only 1 metre per hour, and applied to aerobic organisms such a method will not result in granulation. While under anaerobic conditions the clarification step merely serves to separate organisms and treated water, is also the clarification step in the method according to the invention of essential importance for the induction of granulation.
The invention will now be explained with reference to an exemplary embodiment and with reference to the drawing, in which Fig.- 1 is a graphical representation of the carbon dioxide percentage in gas discharged from a bubbling-bed reactor during a cycle of the method according to the invention; and
Fig. 2 is an illustration of a granule composed of aerobic organisms. Example l
A bubbling-bed reactor (2.5 litres; height/diameter 20) was fed with 1.25 litres of a solution serving as model for waste water, comprising 8.7 mM ethanol, 5 mM ammonium chloride, 4.7 mM potassium phosphate, 2.4 mM magnesium sulphate, 0.48 calcium chloride and per litre solution 1.5 ml of a standard solution of trace elements. The solution was inoculated with aerobically active sludge from a water treatment plant . The model waste water in the bubbling bed reactor was subjected to a cyclic treatment at pH 6-8 and a temperature of 20°C. The treatment consisted of i) aerating for 4 hours at a flow rate of 1.5 litres air per minute (Fig. l shows the carbon dioxide percentage in the gas discharged from a bubbling-bed reac- tor during this phase. This percentage is a measure of the conversion of the ethanol) , ii) the one-minute stoppage of aeration, and iii) draining model waste water from the bubbling-bed reactor at the half-way point of the column of liquid. Any biomass present during draining in the top half of the solution, was discharged together with the effluent. Finally, iv) the bubbling-bed reactor was replenished with a volume of model waste water equal to that of the discharged effluent . The cycle was then resumed with four hours aeration of the solution. Fig. 2 shows the granules comprised of aerobic microorganisms, obtained by the method according to the invention. The average size is 3 mm.

Claims

1. A method of acquiring granular growth of a microorganism in a reactor containing a liquid phase which comprises a substrate, wherein in a first step said substrate is converted by the microorganism resulting in the formation of and growth on a phase comprising the organism while the liquid phase is being mixed, in a sec- ond step mixing in the reactor is stopped to allow part of the solid phase to settle, and in a third step the reactor is partly emptied by discharging the top part of the reactor contents, which reactor is subsequently replenished with substrate-comprising liquid, to repeat steps 1 to 3 , characterized in that the microorganism is an aerobic microorganism, in that at least during the first step a third phase is present, which third phase comprises oxygen-containing gas being fed to the reactor during the first step while the contents of the reactor are kept in turbulence, and in that settling occurs in the second step and takes less time than the height of liquid in the reactor at the end of the first step divided by a settling velocity of at least 5 metres per hour.
2. A method according to claim 1, characterized in that the compound to be converted is fed in pulses.
3. A method according to one of the preceding claims, characterized in that the conversion is a nitrifi- cation-denitrification-conversion in which the oxygen-containing gas is only supplied during the first part of the first step for the completion of the nitrification, and that the reactor operates during the remainder of the first step under substantially anaerobic conditions for the completion of the denitrification.
4. A method according to one of the preceding claims, characterized in that mixing is carried out by feeding oxygen-containing gas.
5. A method according to one of the preceding claims, characterized in that the contents of the reactor are substantially continuously mixed during the first step.
6. A method according to one of the preceding claims, characterized in that when the reactor is started up, a carrier particle is present for the organism.
7. A method according to claim 6 , characterized in that the carrier particle comprises a mycelium-forming fungus .
8. A method according to claim 1 or 2, characterized in that settling occurs in the second step, taking less time than the height of liquid in the reactor at the end of the first step divided by a settling velocity of at least 10 metres per hour, and preferably at least 15 metres per hour.
9. A method according to claim 3 , characterized in that settling occurs taking less time than the height of liquid in the reactor at the end of the first step divided by a settling velocity of at least 15 metres per hour.
PCT/NL1998/000100 1997-02-21 1998-02-19 Method for acquiring grain-shaped growth of a microorganism in a reactor WO1998037027A1 (en)

Priority Applications (5)

Application Number Priority Date Filing Date Title
EP98905876A EP0964831B1 (en) 1997-02-21 1998-02-19 Method for acquiring grain-shaped growth of a microorganism in a reactor
US09/367,985 US6566119B1 (en) 1997-02-21 1998-02-19 Method for acquiring grain-shaped growth of a microorganism in a reactor
DE1998607659 DE69807659T2 (en) 1997-02-21 1998-02-19 METHOD FOR GENERATING GRAIN-SHAPED MICRO-ORGANISM GROWTH IN A REACTOR
AT98905876T ATE223356T1 (en) 1997-02-21 1998-02-19 METHOD FOR GENERATING GRAIN-SHAPED MICROORGANISM GROWTH IN A REACTOR
DK98905876T DK0964831T3 (en) 1997-02-21 1998-02-19 Process for producing granular growth of a microorganism in a reaction vessel

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
NL1005345 1997-02-21
NL1005345A NL1005345C2 (en) 1997-02-21 1997-02-21 Method for obtaining granular growth of a microorganism in a reactor.

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WO1998037027A1 true WO1998037027A1 (en) 1998-08-27

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US (1) US6566119B1 (en)
EP (1) EP0964831B1 (en)
AT (1) ATE223356T1 (en)
DE (1) DE69807659T2 (en)
DK (1) DK0964831T3 (en)
ES (1) ES2182270T3 (en)
NL (1) NL1005345C2 (en)
PT (1) PT964831E (en)
WO (1) WO1998037027A1 (en)

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JP2003033796A (en) * 2001-07-26 2003-02-04 Kurita Water Ind Ltd Biological denitration method
WO2003070649A1 (en) * 2002-02-22 2003-08-28 Sut Seraya Pte Ltd Aerobic biomass granules for waste water treatment
JP2007136363A (en) * 2005-11-18 2007-06-07 Sumitomo Heavy Ind Ltd Granular microbial sludge generation method
JP2007136367A (en) * 2005-11-18 2007-06-07 Sumitomo Heavy Ind Ltd Biological wastewater treatment apparatus and biological wastewater treatment method
JP2007136365A (en) * 2005-11-18 2007-06-07 Sumitomo Heavy Ind Ltd Method for producing granular microbe sludge
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ATE223356T1 (en) 2002-09-15
DE69807659D1 (en) 2002-10-10
US6566119B1 (en) 2003-05-20
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NL1005345C2 (en) 1998-08-24

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